中国组织工程研究 ›› 2013, Vol. 17 ›› Issue (36): 6388-6395.doi: 10.3969/j.issn.2095-4344.2013.36.002
• 骨髓干细胞 bone marrow stem cells • 上一篇 下一篇
骨髓间充质干细胞的分化与静水压力刺激强度和持续时间
何 川1，2，梁 静2，邓廉夫2，冯建民1
- 1上海交通大学医学院附属瑞金医院骨科，上海市 200025；2上海市伤骨科研究所，上海市 200025
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收稿日期:2013-02-19修回日期:2013-03-20出版日期:2013-09-03发布日期:2013-09-03 -
通讯作者:邓廉夫，博士，研究员，教授。上海市伤骨科研究所，上海市 200025 lfdeng@msn.com -
作者简介:何川☆，男，1971年生，四川省泸州市人，汉族，2005年上海第二医科大学毕业，博士，副主任医师，主要从事关节外科、人工关节的研究。 drhechuan@sina.com -
基金资助:国家自然科学基金项目(81071473)*；上海交通大学医学院自然科学研究基金项目(2008XJ038)*
Correlation between magnitude and duration of hydrostatic pressure and the differentiation of human bone marrow mesenchymal stem cells
He Chuan1, 2, Liang Jing2, Deng Lian-fu2, Feng Jian-min1
- 1 Department of Orthopedics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China; 2 Shanghai Institute of Traumatology and Orthopaedics, Shanghai 200025, China
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Received:2013-02-19Revised:2013-03-20Online:2013-09-03Published:2013-09-03 -
Contact:Deng Lian-fu, M.D., Researcher, Professor, Shanghai Institute of Traumatology and Orthopaedics, Shanghai 200025, China lfdeng@msn.com -
About author:He Chuan☆, M.D., Associate chief physician, Department of Orthopedics, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China; Shanghai Institute of Traumatology and Orthopaedics, Shanghai 200025, China drhechuan@sina.com -
Supported by:National Natural Science Foundation of China, No. 81071473*; Natural Science Foundation of Shanghai Jiao Tong University School of Medicine, No. 2008XJ038*
摘要：
背景：力学信号与骨骼系统的生长发育、修复重建和疾病发生发展有密切联系，其对骨髓间充质干细胞的影响和作用机制值得关注。 目的：探讨静水压力刺激对骨髓间充质干细胞分化的影响及相关机制。 方法：①短期实验：将人骨髓间充质干细胞分别接种于正常DMEM培养基、成骨诱导培养基或含细胞外信号调节激酶1/2抑制剂U0126的DMEM培养基中，利用自制压力加载系统对其施加0，40，80 kPa的静水压强1，4 h。②长期实验：将人骨髓间充质干细胞分别接种于正常DMEM培养基或成骨诱导培养基，施加40 kPa的静水压强4 h/d，持续14 d，以不施加静水压的细胞作对照。 结果与结论：实时定量反转录PCR结果显示，经成骨诱导或40 kPa静水压刺激 4 h后，骨髓间充质干细胞内核心结合因子α1、骨钙素mRNA表达增加，过氧化物酶体增殖物活化受体γ2和脂肪酶mRNA表达降低，80 kPa静水压刺激没有出现这一规律；40 kPa静水压的成骨诱导作用可被U0126部分拮抗。组织化学染色显示40 kPa静水压刺激7 d，骨髓间充质干细胞碱性磷酸酶表达和活性均增加；持续刺激14 d，过氧化物酶体增殖物活化受体γ2和脂肪酶mRNA表达增加。说明一定强度和作用时间的静水压刺激可调节骨髓间充质干细胞分化，其作用部分依赖于细胞外信号调节激酶1/2信号途径。
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引用本文
何 川，梁 静，邓廉夫，冯建民. 骨髓间充质干细胞的分化与静水压力刺激强度和持续时间[J]. 中国组织工程研究, 2013, 17(36): 6388-6395.
He Chuan, Liang Jing, Deng Lian-fu, Feng Jian-min. Correlation between magnitude and duration of hydrostatic pressure and the differentiation of human bone marrow mesenchymal stem cells[J]. Chinese Journal of Tissue Engineering Research, 2013, 17(36): 6388-6395.
Morphology of the isolated expanded human bone marrow mesenchymal stem cells
Multipotential of isolated human bone marrow mesenchymal stem cellsThe differentiation potential of the human bone marrow mesenchymal stem cells from three adult donors was observed by culturing the cells under conditions that favorable for adipogenic or osteogenic differentiation. After 1-3 weeks in lineage-specific culture conditions, the expanded cells were highly differentiated (the phenotype of lineage-specific cell types was easily distinguished) without evidence of the other lineages (Figure 3).
Effects of short-time hydrostatic pressure on the differentiation of human bone marrow mesenchymal stem cells Human bone marrow mesenchymal stem cells cultured in Dulbecco’s modified Eagle’s medium and stimulated with 40 kPa pressure for 4 hours showed a significant increasing in the expression of the osteogenic marker gene of core binding factor α1 and osteocalcin, and a significant decreasing in the expressions of the adipogenic marker genes of peroxisome proliferator- activated receptor γ2 and adipsin, when compared with controls (P < 0.05). Human bone marrow mesenchymal stem cells subjected to 80 kPa pressure for 4 hours showed a decreasing in core binding factor α1 expression (P < 0.05). Except for a decreasing in adipsin expression, there were no significant differences between cells cultured in osteogeneic supplemented medium in the presence of hydrostatic pressure and control cells. When extracellular signal-regulated kinase 1/2 activity was blocked by U0126, a significant decreasing in core binding factor α1 expression was observed in cells pressurized with 40 kPa for 4 hours, although core binding factor α1 expression was still higher than that in the non-pressurized control cells. Furthermore, pressurized cells with extracellular signal-regulated kinase 1/2 inhibition showed a decreasing in osteocalcin expression, reaching levels of expression showed no statistically significant difference from that of nonpressurized control cells. In contrast, pressurized cells with extracellular signal-regulated kinase 1/2 inhibition showed a significant increasing of peroxisome proliferator-activated receptor γ2 expression and adipsin expression (Figure 4).
Effects of long-term hydrostatic pressure on the differentiation of human bone marrow mesenchymal stem cells (Figure 5) Hydrostatic pressure of 40 kPa for 4 hours per day could promote the expression of the osteogenic marker genes core binding factor α1 and osteocalcin after 7 days. Interestingly, the same hydrostatic pressure conditions continued for up to 14 days had a very different effect on cells, it could increase the expressions of adipogenic marker genes of peroxisome proliferator-activated receptor γ2 and adipsin. There was no synergistic effect under hydrostatic pressure and osteogenic induction.
Qualitative and quantitative analysis of alkaline phosphatase after long-term hydrostatic pressure stressHydrostatic pressure or osteogenic supplemented medium alone could increase the activity and expression of alkaline phosphatase in human bone marrow mesenchymal stem cells, but this effect was not observed when two stimulus combined (Figure 6).
Cell morphology was then observed, and pictures were taken under phase contrast microscope. After 2-4 days in culture, a heterogeneous population of mononuclear cells (with adherence capacity) including fibroblast-like spindle-shape cells, monocytes, macrophages, endothelial cells, and multinucleated osteoclasts attached to the surface of the dishes. As reported by others[1-5, 13], serial observation of cell shape and size showed that only mesecnhymal stem cell-derived fibroblast-like cells could be proliferated to form colony-forming unit fibroblast colonies (Figure 2). Small fibroblastoid colonies formed at 5-7 days. At the end of the incubation period (13 days), colonies were clearly formed (Figure 2).
Multipotential of isolated human bone marrow mesenchymal stem cellsThe differentiation potential of the human bone marrow mesenchymal stem cells from three adult donors was observed by culturing the cells under conditions that favorable for adipogenic or osteogenic differentiation. After 1-3 weeks in lineage-specific culture conditions, the expanded cells were highly differentiated (the phenotype of lineage-specific cell types was easily distinguished) without evidence of the other lineages (Figure 3).
Effects of short-time hydrostatic pressure on the differentiation of human bone marrow mesenchymal stem cells Human bone marrow mesenchymal stem cells cultured in Dulbecco’s modified Eagle’s medium and stimulated with 40 kPa pressure for 4 hours showed a significant increasing in the expression of the osteogenic marker gene of core binding factor α1 and osteocalcin, and a significant decreasing in the expressions of the adipogenic marker genes of peroxisome proliferator- activated receptor γ2 and adipsin, when compared with controls (P < 0.05). Human bone marrow mesenchymal stem cells subjected to 80 kPa pressure for 4 hours showed a decreasing in core binding factor α1 expression (P < 0.05). Except for a decreasing in adipsin expression, there were no significant differences between cells cultured in osteogeneic supplemented medium in the presence of hydrostatic pressure and control cells. When extracellular signal-regulated kinase 1/2 activity was blocked by U0126, a significant decreasing in core binding factor α1 expression was observed in cells pressurized with 40 kPa for 4 hours, although core binding factor α1 expression was still higher than that in the non-pressurized control cells. Furthermore, pressurized cells with extracellular signal-regulated kinase 1/2 inhibition showed a decreasing in osteocalcin expression, reaching levels of expression showed no statistically significant difference from that of nonpressurized control cells. In contrast, pressurized cells with extracellular signal-regulated kinase 1/2 inhibition showed a significant increasing of peroxisome proliferator-activated receptor γ2 expression and adipsin expression (Figure 4).
Effects of long-term hydrostatic pressure on the differentiation of human bone marrow mesenchymal stem cells (Figure 5) Hydrostatic pressure of 40 kPa for 4 hours per day could promote the expression of the osteogenic marker genes core binding factor α1 and osteocalcin after 7 days. Interestingly, the same hydrostatic pressure conditions continued for up to 14 days had a very different effect on cells, it could increase the expressions of adipogenic marker genes of peroxisome proliferator-activated receptor γ2 and adipsin. There was no synergistic effect under hydrostatic pressure and osteogenic induction.
Qualitative and quantitative analysis of alkaline phosphatase after long-term hydrostatic pressure stressHydrostatic pressure or osteogenic supplemented medium alone could increase the activity and expression of alkaline phosphatase in human bone marrow mesenchymal stem cells, but this effect was not observed when two stimulus combined (Figure 6).
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Of all tissues in the body, bone is probably the tissue that most closely associated with mechanical forces. Mechanical stress is known to be one of the important factors in the regulation of development, remodeling and pathology of bone. The effects of mechanical stress on cells are dependent on the magnitude, duration, and frequency of mechanical stress[1]. Bone cells, especially osteoblasts and osteocytes, respond to mechanical stimulation with the changes in cellular reactions, which in turn regulate bone formation and remodeling[2]. It has been proposed that mature cells such as osteocytes and osteoblasts are responsible for sensing and responding to mechanical stimulus. Bone marrow
In the bone, the transmission of mechanical signals is through fluid flow, which influences cells in the form of hydrostatic pressure and fluid shear stress[8]. Under hydrostatic pressure, cells equally bear load in all directions while under cyclic strain they are subjected to biaxial or multi-axial forces and deform along the force vector. Thus, these two different mechanical signals should trigger different molecular mechanisms within the cells. To test this hypothesis, we used a customized pressure culture system and determined the effect of hydrostatic pressure on the osteogenic and adipogenic differentiation of human bone marrow mesenchymal stem cells.
mesenchymal stem cells, the progenitors that give rise to osteocytes and osteoblasts, also have been shown to be responsive to mechanical stress[3-5].
Although little is known about the molecular mechanisms by which soluble and mechanical signals regulate function of mesenchymal stem cells, Jaiswal et al [6] showed that the commitment of human bone marrow mesenchymal stem cells to the osteogenic and adipogenic lineages in vitro involves signaling by mitogen-activated protein kinase pathways. In particular, they found that induction of human bone marrow mesenchymal stem cells differentiation to osteoblasts by dexamethasone, ascorbic acid, and glycerophosphate is regulated by the extracellular signal-regulated kinase signaling pathway. Furthermore, blocking the extracellular signal-regulated kinase 1/2 pathway can inhibit osteogenic differentiation of human bone marrow mesenchymal stem cells and lead to adipogenesis. Simmons et al [7] reported that cyclic strain can enhance matrix mineralization by human bone marrow mesenchymal stem cells via the extracellular signal-regulated kinase 1/2 signaling pathway. Thus, mitogen-activated protein kinase pathways, which are generally activated by growth factors/cytokines and integrin-mediated cell adhesion, may play critical roles in directing mesenchymal stem cell commitment by mechanical signals.
Design: Cytology contrast study.
Time and setting: The experiment was completed in the Shanghai Institute of Traumatology and Orthopaedics from June 2009 to March 2011.
Materials
The written informed consent was obtained from patients, and then the bone marrow was obtained from human donors undergoing surgery in Shanghai Ruijin Hospital from 2009 to 2010. Bone marrow samples (10-20 mL) were procured from proximal femurs according to the method described previously[9-10]. Six male patients were included in this study, two patients (32 and 38 years old, respectively) with closed diaphyseal femoral fractures and four patients (48, 55, 63 and 66 years old, respectively) with hip osteoarthritis. The mean donor age was 50.3 years (range 32-66 years). Their clinical characteristics and the inclusion criteria were described in our previous report[11].
Experimental reagents and apparatus are as follows:
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Methods
Cell culture conditions
Mesenchymal stem cells were isolated and cultured as previously described[9-11]. In brief, mesenchymal stem cells were isolated by density separation using Histopaque-1077 and subsequent plastic adherence. The cells were purified using a density gradient solution and cultured in vitro using a standard medium (Dulbecco’s modified Eagle’s medium/Ham’s F-12 1:1 with L-glutamine, 10% human serum, 5 µg/mL ascorbic acid). Culture dishes were incubated at 37 ℃ in a humidified atmosphere of 5% carbon dioxide and 95% air. Culture medium was changed after the first four days and thereafter every two days. After reaching confluence (usually about three to four weeks), cells were released with 0.05% trypsin. For first passage culture (P1), an equal number of cells were plated in 75-cm2 flasks. Cells were passaged at 70%-80% confluence and passages 3-7 were used for experiments. Cells capacity to differentiate into the osteogenic and adipogenic, and chondrogenic lineage by addition of appropriate media was also confirmed.
Application of hydrostatic pressure in cultured cells
Cells were placed on 6-well culture dishes. The cells were allowed to attach and spread for 24 hours, and then placed into the pressure vessel. In the short-term loading study, cells were divided into three groups: (1) cells cultured in a standard medium (Dulbecco’s modified Eagle’s medium); (2) cells cultured in osteogenic supplemented medium (serum-containing medium supplemented with 50 mg/mL ascorbic acid, 10 mmol/L β-glycerophosphate, and 0.1 mmol/L dexamethasone); (3) cells cultured in standard Dulbecco’s modified Eagle’s medium containing
10 mmol/L U0126, which blocks the extracellular signal-regulated kinase 1/2 pathway. Cells were then immediately subjected for mechanical stimulation. Using a customized air pressure vessel (Figure 1), a sustained hydrostatic pressure of 40 or 80 kPa was applied to the cells for 1 or 4 hours. In the long-term study, a sustained hydrostatic pressure of 40 kPa was applied to human bone marrow mesenchymal stem cells (cultured in 10% Dulbecco’s modified Eagle’s medium or osteogenic supplemented medium) for 4 hours every day and lasted for 14 days. Control culture plates were setup in identical conditions, except that no pressure was applied.
10 mmol/L U0126, which blocks the extracellular signal-regulated kinase 1/2 pathway. Cells were then immediately subjected for mechanical stimulation. Using a customized air pressure vessel (Figure 1), a sustained hydrostatic pressure of 40 or 80 kPa was applied to the cells for 1 or 4 hours. In the long-term study, a sustained hydrostatic pressure of 40 kPa was applied to human bone marrow mesenchymal stem cells (cultured in 10% Dulbecco’s modified Eagle’s medium or osteogenic supplemented medium) for 4 hours every day and lasted for 14 days. Control culture plates were setup in identical conditions, except that no pressure was applied.
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Osteogenic and adipogenic differentiation related mRNA expression detected by real-time quantitative reverse transcriptase PCR
Total cellular RNA was extracted from mesenchymal stem cells using Trizol according to the single step acid-phenol guanidinium method[12]. cDNA was synthesized from the RNA using these cells. Real-time quantitative reverse transcriptase-PCR was performed by using an RG-3000 Sequence Detection System. The expression level of each target gene was normalized to the reference gene glyceraldehyde phosphate dehydrogenase and the resulting data were expressed as a ratio of the control. Data were collected with instrument spectral compensations by the applied biosystems of sequence detection system 2.1 software, and analyzed using the threshold cycle number (Ct) relative quantification method. Primers and probes are as follows: glyceraldehyde phosphate dehydrogenase: sense was 5’-ACC ACA GTC CAT GCC ATC AC-3’, antisense was 5’-TCC ACC ACC CTG TTG CTG TA-3’; core binding factor alpha 1: sense was 5’-CCG CCC CAC GAC AAC CGC ACC AT-3’, antisense was 5’-GCT CTG TGA TAG GTA GCT AC-3’; osteocalcin: sense was 5’-CTC ACA CTC CTC GCC CTA T-3’, antisense was 5’-CAG CCA ACT CGT CAC AGT C-3’; peroxisome proliferator-activated receptor γ2: sense was 5’-TTG AAT GTC GTG TCT GTG GAG AT-3’, antisensewas 5’-CAG CGG GAA GGA CTT TAT GTA TG-3’; adipsin: sense was 5’-GCT GGG GCA TAG TCA ACC AC-3’, antisense was 5’-GTA GAT CCC GGG CTT CTT GC-3’.
Qualitative and quantitative analysis of alkaline phosphatase
Human bone marrow mesenchymal stem cells were pressurized for up to 7 or 14 days while maintained in Dulbecco’s modified Eagle’s medium or osteogenic Supplemented medium. Alkaline phosphatase activity, an early but nonspecific marker of osteogenesis, and mineral deposition, a late differentiation marker, were assessed qualitatively by histochemical staining. Cells were fixed in 70% ethanol and stained for alkaline phosphatase activity using alkaline phosphatase activity kit. To quantitatively measure alkaline phosphatase activity, cell layers were washed twice with PBS and then harvested in a passive lysis buffer. The lysate was sonicated and centrifuged. The supernatant was tested for alkaline phosphatase activity by incubation with 50 mmol/L p-nitrophenyl phosphate in an assay buffer (100 mmol/L glycine,
1 mmol/L MgCl2, pH 10.5) at 37 ℃ for 10-20 minutes. The absorbance was measured with an ultraviolet spectrophotometer DU640 at 405 nm and converted to p-nitrophenol concentration based on standard solutions prepared in parallel. To determine the amount of DNA in each well, the cell nuclei were disrupted by addition of lysis buffer (0.025 mol/L tris-HCl, 0.4 mol/L NaCl, 0.5% SDS, pH 7.4), followed by sonication and centrifugation. The DNA content was determined from the supernatant using the Hoescht 33258 assay and standard solutions of calf thymus DNA prepared in parallel.
1 mmol/L MgCl2, pH 10.5) at 37 ℃ for 10-20 minutes. The absorbance was measured with an ultraviolet spectrophotometer DU640 at 405 nm and converted to p-nitrophenol concentration based on standard solutions prepared in parallel. To determine the amount of DNA in each well, the cell nuclei were disrupted by addition of lysis buffer (0.025 mol/L tris-HCl, 0.4 mol/L NaCl, 0.5% SDS, pH 7.4), followed by sonication and centrifugation. The DNA content was determined from the supernatant using the Hoescht 33258 assay and standard solutions of calf thymus DNA prepared in parallel.
Main outcome measures
Osteogenic and adipogenic differentiation of human bone marrow mesenchymal stem cells under different hydrostatic pressures stimulation, and the effect of extracellular signal-regulated kinase 1/2 on the differentiation were measured.
Statistical analysis
Independent experiments were performed at least four times in duplicate cultures (unless otherwise noted). Data were expressed as mean±SEM and analyzed for levels of significance with SPSS 11.5 using the Student’s t-test (one sample t-test and paired t-test, where appropriate). P value < 0.05 was considered as statistical significant.
Mechanical load, such as gravity and body posture, produces biomechanical stimulations in the bone tissue, and plays an important role in the process of bone development, growth, modeling and remodeling. The differentiation of mesencnymal stem cells and the progenitors of multiple cellular lineages in bone, may decide the quality, quantity, and structure of the new bone formation. Mesenchymal stem cells have been proven to be responsive to mechanical signals such as hydrostatic pressure[13] and to shear stress induced by fluid flow[14] in their environment. High intraosseous pressure, suggesting high hydrostatic pressure in bone, is usually occurs in the pathology of some bone diseases, such as osteonecrosis, osteoarthritis, and osteomyelitis. We demonstrated herein that hydrostatic pressure can influence the osteogenic and adipogenic differentiation of human bone marrow mesenchymal stem cells in vitro.
In conclusion, our results suggest that hydrostatic pressure can influence the differentiation of human bone marrow mesenchymal stem cells. These cells are sensitive to the patterns, duration, and magnitude of mechanical stimulations which trigger different mechanical signals.
It reported that cyclical stress (stretching) could promote the osteogenic differentiation of human bone marrow mesenchymal stem cells, especially matrix mineralization[7]. Hydrostatic pressure and stretching may produce different mechanical signals. Stretching makes cells deformed along one or two directions, while hydrostatic pressure exerts a mechanical load on cells from all directions equally, and cells do not deform, except for a decrease in height. Simmons et al reported that stretching could stimulate an increase in matrix mineralization of human bone marrow mesenchymal stem cells cultured in osteogenic supplemented medium. We showed that hydrostatic pressure could inhibit osteogenic differentiation of mesenchymal stem cells stimulated by osteogenic supplemented medium. But, in certain condition, hydrostatic compression alone can promote osteogenic differentiation of human bone marrow mesenchymal stem cells, and long-term pressurization can promote adipogenic differentiation. These results show that the effects of hydrostatic pressure and stretching on mesenchymal stem cells are different.
In current study, we found that the effects of hydrostatic pressure on the differentiation of mesenchymal stem cells are dependent on the magnitude and duration of stress. Relative lower pressure (40 kPa) and short-term (7 days) may promote mesenchymal stem cells to differentiate into the osteogenic lineage, and higher pressure (80 kPa) and long-term (14 days) seemed to promote the adipogenic differentiation of mesenchymal stem cells. We demonstrated in previous study that the cytoskeletal structure of mesenchymal stem cells would be reorganized with hydrostatic pressure loading. It is interesting that the pattern of the cytoskeletal structure reorganization and rearrangement are related to the magnitude and duration of stress. The actin filaments of the strained bone marrow were more flimsy and tenuous than un-treated osteoblasts. Unlike the normal distribution of bundles or membrane-like of the control group, they were arranged without direction. But after stained for 4 hours, cells showed an arrangement more close to the unstained cells than that after stained for 1 hour. The flow-induced changes in cytoskeletal stresses are also found to be dynamic, and the cytoskeletal changes are accompanied by reorganization of the actin cytoskeleton from parallel F-actin bundles to peripheral bundles[15]. Steward et al recently reported the cytoskeleton mediated regulating chondrogenesis of mesenchymal stem cells under hydrostatic pressure that similar to our results[16]. They found that intermediate filaments also appear to play a role in the mechanotransduction of hydrostatic pressure, as only vimentin organisation adapted in response to this mechanical stimulus[16]. It may explain the effects of hydrostatic pressure on the differentiation of mesenchymal stem cells and the changes of cytoskeletal structure of mesenchymal stem cells with the magnitude and duration of hydrostatic pressure loading.
Strech-induced osteoblastic genes expression is mainly mediated by extracellular signal-regulated kinase 1/2 signaling. Ziros et al [17] reported that low levels of mechanical deformation (stretching) of human osteoblastic cells could directly up-regulate the expression and DNA binding activity of the osteoblast-specific transcription factor core binding factor α (core-binding factor), which plays a fundamental role in differentiation and function of osteoblasts. They found that this effect seems to be well tuned by the stretch-mediated induction of distinct mitogen-activated protein kinase cascades. Simmons et al [7] also demonstrated that strech-induced mineralization was largely mediated by the extracellular signal-regulated kinase 1/2 pathway. Sustained hydrostatic pressure can induce protein tyrosine phosphorylation, activation of extracellular signal-regulated kinase 1/2, and can enhance expression of c-Fos in both time- and magnitude-dependent manners. In our previous study, the U0126 interrupting the rearrangement phenomenon of bone marrow mesenchymal stem cells under hydraulic pressure suggests that the extracellular signal-regulated kinase 1/2 signaling pathway plays an important role in the process[11] when compared with other reports[13, 18]. In current study, however, we found that compression- induced osteogenic differentiation is only partly mediated by the extracellular signal-regulated kinase 1/2 pathway. When the extracellular signal-regulated kinase 1/2 pathway was blocked with U0126, an extracellular signal-regulated kinase 1/2 inhibitor, core binding factor α expression in human bone marrow mesenchymal stem cells stimulated with 40 kPa hydrostatic pressure for 4 hours was 6 times higher than that in un-pressurized control cells. Thus, pressure-induced osteoblastic differentiation in human bone marrow mesenchymal stem cells seems to involve other signaling pathways.
1力学信号是肌肉骨骼组织生长发育的重要调控因素，在细胞水平的修复过程中力学信号也起重要的调控作用。 2骨髓间充质干细胞成骨/成脂分化调控涉及许多骨科疾患的病理演变过程，实验重点研究压应力信号对其影响。 3 实验结果发现，40 kPa静水压可以调节骨髓间充质干细胞分化，其作用部分依赖于细胞外信号调节激酶1/2信号途径。
Mechanical load, such as gravity and body posture, produces biomechanical stimulations in the bone tissue, and plays an important role in the process of bone development, growth, modeling and remodeling. The differentiation of mesencnymal stem cells and the progenitors of multiple cellular lineages in bone, may decide the quality, quantity, and structure of the new bone formation. Mesenchymal stem cells have been proven to be responsive to mechanical signals such as hydrostatic pressure and to shear stress induced by fluid flow in their environment. High intraosseous pressure, suggesting high hydrostatic pressure in bone, is usually occurs in the pathology of some bone diseases, such as osteonecrosis, osteoarthritis, and osteomyelitis. We demonstrated herein that hydrostatic pressure can influence the osteogenic and adipogenic differentiation of human bone marrow mesenchymal stem cells in vitro.
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